1. The Field of the Invention
The present invention generally relates to the isolation of certain toxins from spider venoms and the use of those toxins as inhibitors of the functions of ion channels and neurotransmitter receptors. In particular, the present invention relates to spider venom toxins and their use as blockers of calcium channels and excitatory amino acid receptors in the cardiovascular, central nervous, and neuromuscular systems of organisms, including humans.
2. The Background of the Invention
Movement of calcium ions across cell membranes is a critically important event in the normal functioning of excitable tissues such as vascular smooth muscle, cardiac muscle, and the central nervous system. Influx of calcium ions through specialized channels in the cell membranes regulates release of substances such as hormones and neurotransmitters.
The movement of calcium ions also regulates contraction of heart muscle and of vascular smooth muscle in the wall of blood vessels. Abnormal influx of calcium ions has been reported to play a role in the pathogenesis of various cardiovascular disorders (e.g., anoxic/ischemic heart disease), and drugs capable of blocking the movement of calcium through calcium channels have been used for treatment of cardiac arrhythmias, coronary artery disease, and cardiomyopathy.
The currently used drugs, however, have non-specific physiological effects and varying tissue specificities that can lead to undesirable side-effects in patients. Moreover, there are several known subtypes of calcium channel with varying physiological action and no drug that specifically blocks certain of these subtypes is known.
In the nervous system, calcium influx into the presynaptic nerve terminal via calcium channels is a necessary prerequisite for the release of chemical neurotransmitter at synapses and thus for the proper functioning of these synapses. Lowering of the extracellular calcium concentration is routinely used by neurophysiologists to reduce or abolish synaptic transmission in isolated pieces of nervous tissue.
It has not been possible, however, to specifically affect synaptic transmission in vivo in the central nervous system ("CNS") by manipulating the function of neuronal calcium channels. With the exception of the omega-conotoxin recently isolated from the venom of the marine snail Conus geographus, no drug with sufficiently specific or potent effects on CNS calcium channels is known.
Abnormal influx of calcium is thought to be very important in the pathogenesis of several CNS disorders, including anoxic/ischemic (stroke) damage, epilepsy, and the neuronal death associated with chronic epilepsy. Again, the paucity of chemical agents that potently and specifically block CNS calcium channels has prevented the development of an effective drug therapy for these prevalent neurological problems.
In addition, excitatory amino acids ("EAA"), most notably glutamate and aspartate, are the predominant excitatory neurotransmitter in the vertebrate (including human) CNS. As such, they play a fundamental role in the many functions of the normal nervous system. EAA's are released from presynaptic nerve terminals and, after diffusing across the synaptic cleft, contact special EAA receptor molecules in the postsynaptic cell membrane. These receptors indirectly influence the flow of various ions across the cell membrane and thus contribute to production of an electrical response to the chemical message delivered by EAA neurotransmitter molecules. A number of common and very serious neurological problems involve abnormal function of EAA synapses. These include epilepsy, several degenerative disorders such as Huntington's disease, and neuronal death following stroke.
Unfortunately, there are very few chemical agents which are potent and selective blockers (that is "antagonists") of EAA receptors. This has severely hampered research on the normal function of EAA's and limited therapeutic approaches to disorders involving EAA's.
One notable limitation of the currently available EAA receptor antagonists is the lack of any drugs with very high affinity for the receptor. A drug with high affinity for the receptor could be expected to produce irreversible blockade of synaptic transmission. When labeled with some tracer molecule, such a drug would provided a reliable way of tagging receptors to permit measurement of their number and distribution within cells and tissues. These features would have very valuable consequences for research on EAA neurotransmission and for the development of therapeutic agents to treat EAA dysfunction in humans and animals.
Arthropod animals, including insects, and certain parasitic worms use EAA's as a major chemical neurotransmitter at their neuromuscular junction and in their CNS. Because of the damage done by insect pests and the prevalence of parasitic worm infections in animals and humans in many countries, there is a constant need for potent and specific new pesticides and anthelmintic drugs that are non-toxic to humans, pets, and farm animals.
As described above, it would be a very considerable improvement in the art if it were possible to develop chemical agents that specifically and potentially block calcium channel function in the CNS, cardiac muscle, and vascular smooth muscle. In particular, it would be an advancement in the art to provide a specific blocker for specific subtypes of calcium channel. Similarly, it would be an advancement in the art to provide a specific blocker of calcium channels in the CNS.
It would be a further significant advancement in the art if chemical compositions could be found that specifically bind to and block the function of EAA in the CNS of vertebrates (including humans) and in the nervous systems of invertebrates. It would be a further advancement in the art to provide calcium channel blockers and EAA receptor blockers whose actions were selectively reversible or irreversible for use in clinical settings or as research tools. It would be an additional advancement in the art to provide calcium channel blockers and EAA receptor blockers that were acceptable for use as insecticides and anthelmintics.
Such chemical compositions and methods for their use are disclosed and claimed below.